Chiral synthesis of anthracyclines from substituted anthraquinones

ABSTRACT

Anthracycline glycosides of formula (I): ##STR1## wherein R 1  and R 2  are independently selected from hydrogen, methyl, ethyl or methoxy and pharmaceutically acceptable acid addition salts thereof, which are useful in treatment of human cancer, are prepared by a stereospecific condensation of a derivative of daunosamine of general formula (II): ##STR2## wherein R 3  is an alkyl, perfluoroalkyl or aryl group and R 4 , R 5 , R 6  and R 7  are independently selected from alkyl, aryl or aralkyl having up to ten carbon atoms, with a quinone quinizarine epoxide of general formula (III) ##STR3## and conversion of the product obtained to the anthracycline glycoside of formula (I) or salt thereof.

DESCRIPTION

This invention relates to a multistep method for the stereospecificsynthesis of anthracyclines of general formula (I): ##STR4## wherein R₁and R₂ are independently selected from hydrogen, methyl, ethyl andmethoxy, and pharmaceutically acceptable acid addition salts thereof.

Anthracyclines of general formula (I) exhibit a strong antitumoractivity. 4-Demethoxydaunomycin for example (I, R₁ ═R₂ ═H;) has a higheranti-leukemic activity than daunomycin and adriamycin and is alsoeffective by oral administration. Many synthetic approaches to theaglycones anthracyclinones have been disclosed. However, the methods aregenerally inadequate to prepare them in optically pure form. Theinvention provides a sequence of sterospecific reactions leading to thedesired anthracyclines (I) in the optically pure form as required fortherapeutic use.

Accordingly, the present invention provides a stereospecific process forthe preparation of an anthracycline glycoside of general formula (I):##STR5## wherein R₁ and R₂ are independently selected from hydrogen,methyl, ethyl or methoxy, or a pharmaceutically acceptable acid additionsalt thereof; which process comprises

(i) reacting a daunosamine silyl derivative of general formula (II):##STR6## wherein R₃ is an alkyl, perfluoroalkyl or aryl group and R₄,R₅, R₆ and R₇ are independently selected from alkyl, aryl or aralkylhaving up to ten carbon atoms, with a quinone quinizarine epoxide ofgeneral formula (III): ##STR7## in an inert organic solvent;

(ii) removing the silyl group from the protected tetracyclic derivativeof general formula (VIII): ##STR8## wherein R₁, R₂, R₃, R₄, R₅, R₆ andR₇ are as above defined, in an inert organic solvent;

(iii) converting the resultant ketone of general formula (IX): ##STR9##wherein R₁, R₂, R₃ and R₄ are as defined above, into the intermediatecompound of general formula (X): ##STR10## wherein R₁, R₂, R₃ and R₄ areas defined above; and

(iv) converting the compound of general formula (X) into ananthracycline glycoside of formula (I) or a pharmaceutically acceptableacid addition salt thereof.

The compounds of formula (II) also form part of the invention.Preferably R₃ represents a C₁ -C₄ alkyl, perfluoromethyl or phenylgroup. Preferably, R₄, R₅, R₆ and R₇ are each independently a C₁ -C₄alkyl, phenyl or benzyl group. More preferably, R₄ is p-nitro-phenyl andR₅, R₆ and R₇ each represent methyl.

A compound of formula (II) is prepared according to the invention by aprocess which comprises

(A) converting a daunosamine of formula (IV): ##STR11## or an acidaddition salt thereof, into a protected derivative of general formula(V): ##STR12## wherein R₃ and R₄ are as defined in above;

(B) selectively deacylating the protected derivative of general formula(V) to give a daunosamine N,O protected derivative of general formula(VI): ##STR13## wherein R₃ and R₄ are as defined above;

(C) reacting the daunosamine N,O protected derivative of formula (VI)with 3-butyn-2-one to give a mixture of the α- and β-anomeric compoundsof general formula (VII) and (VIIA): ##STR14## wherein R₃ and R₄ are asdefined above;

(D) separating the α-anomeric compound of formula (VII) from theβ-anomeric compound of formula (VIIA); and

(E) silylating the α-anomeric compound of formula (VII) so as to obtainthe compound of formula (II).

The general reaction scheme is set forth in the SCHEME below. Itincludes the key steps of synthesising the hitherto, unknown daunosaminederivatives (II) and of condensing this compound with a quinonequinizarine epoxide (III). The latter can be easily prepared fromcommercially available starting material according to known methods(e.g. U.S. Pat. Nos. 4,021,457, 4,070,382, J. Org. Chem. 41, 2296 (1976)and J. Chem. Soc. Chem. Comm. 1981, 478 and the references quotedtherein).

The daunosamine derivative (II) is prepared from a naturally occurringaminosugar daunosamine (IV) or an acid addition salt thereof, preferablythe hydrochloride. For this purpose, compound (IV) is initiallytransformed into the intermediate (V) according well known methods [e.g.T. H. Smith et al., J. Org. Chem. 42, 3653 (1977)]. The intermediate (V)is then selectively deacylated (deacetylated when R₄ is methyl) to givethe daunosamine N,O protected derivative (VI). Such selectivedeacylation is performed typically by dissolving the compound (V) in anorganic solvent such as one chosen from ketones, esters, aliphatic orcycloaliphatic ethers and polyethers. The compound (V) may then betreated with a catalytic amount of a mineral acid, for example by addingat least one equivalent of H₂ O and a catalytic amount of mineral acid.4N aqueous hydrochloric acid may be used. The reaction time generallyfrom 2 to 24 hours and the temperature should be kept at from -10° C. to+50° C.

The next step of the present invention consists in reacting (VI) with3-butyn-2-one. Typically, at least one equivalent of the 3-butyn-2-oneis used. Preferably, the reaction is effected in the presence of acatalytic amount of an aliphatic or cycloaliphatic amine, generally atertiary amine such as trimethylamine, triethylamine, tripropylamine,N-alkylpyrrolidine, N-alkylpiperidine, N-alkylpiperidine,N-alkylmorpholine and tetramethylguanidine. The reaction is normallycarried out in an inert organic solvent, preferably an aromatichydrocarbon such as benzene, toluene or xylene. The reaction may takeplace at about 80° C. for about 1 hour. A mixture of the product (VII)and of its anomer (VIIA) is formed. The desired compound (VII) can beisolated from the mixture by conventional methods such as fractionalcrystallisation or chromatography. The latter is generally preferredgiving (VII) in very pure form. The anomer (VIIA) which can not be usedfor the purpose of the present invention can be transformed back to thestarting material (VI) and can therefore be used for furtherpreparations of (VII).

The compound (VII) is silylated such as to obtain the silyl derivative(II). For the purpose, (VII) is preferably dissolved in an anhydrousinert organic solvent, preferably chosen among aromatic hydrocarbons orchlorinated aromatic or aliphatic hydrocarbons (e.g. CH₂ Cl₂, CHCl₃, C₂H₄ Cl₂, chlorobenzene, benzene or toluene) or their mixtures. Normallyan inert atmosphere is employed. Compound (VII) can be reacted with asilyl derivative of general formula

    SiR.sub.5 R.sub.6 R.sub.7 Y

where R₅, R₆, R₇ are independently selected in the group of alkyl oraryl-alkyl having up to ten carbon atoms and Y represents Cl, Br, I, CN,NHSiR₅ R₆ R₇, ##STR15## CF₃ SO₂ or S-SiR₅ R₆ R₇. Typically at least oneequivalent of the silyl derivative SiR₅ R₆ R₇ Y is used and reaction iscarried out in the presence of at least one equivalent (based on thecompound (II)) of a tertiary aliphatic amine is added. Trimethyl- ortriethyl-amine are preferred. The reaction generally lasts for at least1 h at a temperature between -20° C. and 50° C.

A Diels Alder condensation is then effected between the compound (II)and the quinizarine epoxide (III). The reaction conditions for the DielsAlder condensation may vary in accordance with the substitution patternin the aromatic ring of the quinone quinizarine epoxide and according tothe nature of R₄, R₅, R₆, R₇ on the diene compound (II). Generally,however, the reaction is carried out in an inert organic solvent such anaromatic hydrocarbon or aliphatic or aromatic chlorohydrocarbon at atemperature of from -10° C. to +60° C. The reaction is best run at roomtemperature for from 10 to 24 hours.

Evaporation of the most part of the solvent and addition ofdiethyl-ether can give a precipitate consisting in pure (VIII) which hasnot only the desired tetracyclic skeleton but above all it has thedaunosamyl group at the C₇ in the desired absolute configuration. Theremoval of the silyl group from the compound (VIII) can be performedaccording to conventional methods e.g. hydrolytic cleavage typically inthe presence of strong acids. For this purpose, (VIII) may be dissolvedin an inert solvent selected from ethers, esters, ketones, hydrocarbonsand chloro-carbons and shaken with 0.01 to 2 equivalents of a mineral ororganic acid, eventually dissolved in water.

Transformation of (IX) into (X) can be obtained in a single step byaction of a molar excess of trimethyl silyl iodide. This may be prepared"in situ", from sodium or potassium iodide and trimethyl silyl iodide.This reaction is normally run in an anhydrous solvent. Acetonitrilegives the best result and is the solvent of choice if trimethyl silyliodide has to be prepared "in situ". Other anhydrous solvents can beused without detriment, however. Normally, the reaction is effectedunder a nitrogen atmosphere. Conventional work up such as dilution witha solvent immiscible with water, washing with water and with aqueoussolution of Na₂ S₂ O₃ (to remove the formed elemental iodine), dryingand removal of the solvent gives (X).

The compound (X) is then converted into an anthracycline glycoside (I)or pharmaceutically acceptable salt thereof. Known methods can be used.They are here exemplified to stress the practical relevance of thepresent invention in the synthesis of anthracyclines but they must beconsidered in no way a limitation of the present invention. Thus, aglycoside (I) or salt thereof may be prepared by

(a) converting the compound of general formula (X) into a 9-ethynylderivative of general formula (XI): ##STR16## wherein R₁, R₂, R₃ and R₄are as defined above;

(b) oxidising the 9-ethynyl derivative of formula (XI) to obtain acompound of general formula (XII): ##STR17## wherein R₁, R₂, R₃ and R₄are as defined above;

(C) hydrating the compound of formula (XII) to give an N,O protecteddaunorubicin derivative of general formula (XIII): ##STR18## wherein R₁,R₂, R₃ and R₄ are as defined above;

(d) removing the N- and O-protecting groups from the compound of formula(XIII) to obtain an anthracycline glycoside of general formula (I); and

(e) if desired, converting the anthracycline glycoside of formula (I)into a pharmaceutically acceptable acid addition salt thereof.

The compound (X) for example can react with an ethynyl metal derivativeof general formula M--C.tbd.CH (where M═Li, Na, BrMg, ClMg, 1/2 Mg) togive a derivative (XI). This reaction may be effected at 0° C. in drytetrahydrofuran using 30 equivalents of the ethynyl metal derivative.The resultant compound (XI) can be oxidized e.g. with lead tetraacetateto give the compound (XII). Reaction may take place for 24 hours. Thecompound (XII) can subsequently be hydrated using HgO and aqueous H₂ SO₄to give the N,O protected daunorubicin derivative (XIII). The compound(XII), dissolved in acetone, may be treated with red mercuric oxide and7% aqueous H₂ SO₄ under reflux for 5 minutes.

Removal by hydrolysis of the R₄ CO and R₃ CO groups leads to theglycoside (I). This can be effected by conventional hydrolyticprocedures such as mild alkaline hydrolysis. Thus, the N,O protecteddaunorubicin derivative (XIII) may be treated with 0.1N aqueous NaOH atroom temperature for one hour. Preferably, the resultant anthracyclineglycoside (I) is converted into the hydrochloride salt thereof. This maybe achieved by treating the glycoside (I) with methanolic 0.1N hydrogenchloride and the resultant hydrochloride salt is isolated.

The invention also provides pharmaceutical compositions comprising ananthracycline glycoside of formula (I), or a pharmaceutically acceptableacid addition salt thereof, prepared according to the present inventionand a pharmaceutically acceptable carrier or diluent. Such compositionscontain a therapeutically effective amount of the glycoside or its salt.Conventional formulations, carriers and diluents may be employed.

The following Examples illustrate the invention.

EXAMPLE 1 Preparation of:2,3,6-trideoxy-4-O-p-nitrobenzoyl-3-N-trifluoroacetamido-α-L-lyxohexopyranose(VI)

To a solution of2,3,6-trideoxy-1,4-di-O-p-nitrobenzoyl-3-N-trifluoroacetamido-α-L-lyxohexopyranose(V) (2 g 3.69 mmol.) in acetone (20 ml) were added 3 ml of 4N HCl. After12 h the reaction mixture was concentrated in vacuo, dissolved in ethylacetate (20 ml) and washed with saturated water solution of sodiumbicarbonate and subsequently with water, dried over Na₂ SO₄ andevaporated to afford 1.35 g of (VI) (93% of yield on V)

    ______________________________________                                        H.sup.2 --NMR [300MHz,(CD.sub.3).sub.2 SO]                                                         1.05 (d, 3H, --CH.sub.3)                                                      4.3 (q, 1H, 5 --H)                                                            5.3 (d, 1H, 4 --H)                                                            5.4 (d, 1H, 1 --H)                                                            6.6 (brs, 1H, OH)                                        EI-MS m/e 391                                                                 ______________________________________                                    

EXAMPLE 2 Preparation of(E)-4-(2',3',6'-trideoxy-4'-O-p-nitrobenzoyl-3'-N-trifluoroacetamido-.alpha.-L-lyxohexopyranosyl)oxy-but-3-en-2-one(VII)

To a suspension of (VI) (10 g 25 mmol in toluene (300 ml) at 80° C. wereadded 3-butyn-2-one (2.4 ml 30 mmol and triethylamine (350μp 2.5 mmol.

The reaction was stirred at 80° C. for 3h, allowed to cool to roomtemperature.

Diethyl ether was then added (co. 300 ml).

The formed precipitated was collected, washed with diethyl ether toafford 10.8 g (94% of yield) of a mixture of α,βanomers.

Crystallization of this material from chloroform gave pureβ-anomer(VIIA) (5.3 g).

The mother liquor was purificated by flash-cromatography to give pureα-anomer (VII) (5.4 g) that was crystallized from diethyl ether.

    ______________________________________                                        anomer (VII)                                                                  m.p. 140-142° C.                                                       IR KBr        1730 (C = O)                                                                  1685 (C = O insaturated)                                        UV (ETOH)     237 nm                                                          H.sup.1 --NMR 300 MHZ, (CD.sub.3).sub.2 SO (d, 3H, CH.sub.3)                              1.95 (1H, dd, 2'-Hax)                                                         2.15 (3H, s, CH.sub.3 C = O)                                                  2.45 (1H, dt, 2'-Heq)                                                         4.2 (1H, q, 5'-H)                                                             4.45 (1H, m, 3'-H)                                                            5.35 (1H, d, 4'-H)                                                            5.75 (1H, d, 1'-H = 1.8 Hz)                                                   5.8 (1H, d, 3-H = 12.5 Hz)                                                    7.75 (1H, d, 4-H = 12.5 Hz)                                                   8.22 (2H, d, Arom.)                                                           8.42 (2H, d, Arom.)                                                           9.65 (1H. d. NH)                                                  MS-DCI: m/e 460                                                               β-anomer (VII A)                                                         m.p. 100° C.                                                           H.sup.1 --NMR 300 MHz, (CD.sub.3).sub.2 SO inter alia 5.25 (1H, d, 4'-H)                  5.35 (1H, dd, 1-H 5.8 and 6.2 Hz)                                 ______________________________________                                    

EXAMPLE 3 Preparation of (E)-1-[2',3',6'-trideoxy-4'-O-p-nitrobenzoyl-3'-N-trifluoracetamido-α-L-lyxohexopyranosyl)oxy]-3-trimethylsilyloxybuta-1,3diene(II R₅ ═R₆ ═R₇ ═CH₃)

To a stirred mixture of fused Zinc chloride (200 mg 1.47 mmol) andtriethylamine (5.5 ml 40 mmol) a solution of (VII) (2.3 g 5 mmol) inmethylene chloride (30 ml) was added, followed by trimethylsilylchloride(2.5 ml 20 mmol).

The mixture was stirred at room temperature overnight, then diethylether was added and the mixture was filtered.

After usual work-up a white solid was obtained (2.1 g 78% of yield onVII)

    ______________________________________                                        H.sup.1 --NMR (300 MHz, CDCl.sub.3) 0.26 (s, 9H, CH.sub.3 --Si)                          1.1 (d, 3H, --CH.sub.3)                                                       4.2 (q, 1H, 5'-H)                                                             4.2 (d, 2H, 4-H.sub.2)                                                        5.45 (d, 1H, 1'-H)                                                            5.35 (d, 1H, 4'H)                                                             5.7 (d, 1H, 2-H)                                                              6.8 (d, 1H, 1-H)                                                   MS-DCl: m/e 532                                                               ______________________________________                                    

EXAMPLE 4 Preparation of (VIII) by reaction of epoxytetrone (III) withthe diene (II)

A solution of (III) (1.016 g 4 mmol) and (II) (2.13 g 4 mmol) in drytoluene (80 ml) was stirred at room temperature for 24 h.

After evaporation of solvent, the addition of diethyl ether gave (VIII)as a colourless precipitate (2.3 g 74% of yield)

    ______________________________________                                        m.p. 153-155° C.                                                       H.sup.1 --NMR (300 MHz, CDCl.sub.3)                                                                0.3 (s, 9H, CH.sub.3 --Si)                                                    1.1 (d, 3H, CH.sub.3)                                                         3.9 (t, 1H, 10a-H)                                                            3.2 (dd, 1H, 10a-H)                                                           4.1 (q, 1H, 5'-H)                                                             4.55 (dd, 1H, 7-H)                                                            5.0 (d, 1H, 4'H)                                                              5.1 (d, 1H, 8-H)                                                              5.2 (d, 1H, 1'-H)                                        MS-DCI m/e 786                                                                ______________________________________                                    

EXAMPLE 5 Preparation of (IX)

A solution of (VIII) (1.0 g 1.27 mmol) in freshly distilled THF (15 ml)was treated with 0.1N HCl (1 ml).

When the hydrolysis was completed (TLC analysis) (6 h), the mixture wasdiluted with methylene chloride (50 ml), washed with water and driedover Na₂ SO₄. Evaporation of the organic layer gave (IX) as colourlesssolid (0.7 g. 77% of yield).

    ______________________________________                                        H.sup.1 --NMR (300 MHz, CDCl.sub.3)                                                                 1.1 (d, 3H, CH.sub.3)                                                         3.5 (dd, 1H, 6a-H)                                                            4.8 (q, 1H, 7-H)                                                              5.05 (d, 1H, 4'-H)                                                            5.2 (d, 1H, 4'-H)                                       MS-DCI: m/e 714                                                               ______________________________________                                    

EXAMPLE 6 Preparation of compound (X)

To a solution of sodium iodide (3.9 mmol 0.585 g) in anhydrousacetonitrile (5 ml) trimethylsilylchloride (1.9 mmol 240μl) was addeddropwise under nitrogen. After few minutes, a solution of (IX) (0.93 g1.3 mmol) in anhydrous acetonitrile (3 ml) was slowly added.

After 30' the mixture reaction was tracted with 5N aq sodiumthiosulphate (20 ml), extracted with ethylacetate (three portions of 20ml), and washed with water, dried over Na₂ SO₄ and evaporated to give(X)(0,7 g 77% of yield) as pale yellow solid.

    ______________________________________                                        H.sup.1 --NMR (300 MHz, CDCl.sub.3)                                                             1.2 (d, 3H, CH.sub.3)                                                         3.55 (dd, 1H, 6a-H)                                                           3.82 (dt, 1H, 10a-H)                                                          4.05 (q, 1H, 5'-H)                                                            4.68 (q, 1H, 7-H)                                                             4.84 (d, 1H, 1'-H)                                                            5.3 (d, 1H, 4'-H)                                                             12.8 and 13.7 (s, each 1H, 5 and                                              12-OH)                                                      ______________________________________                                    

EXAMPLE 7 Preparation of 4-demethoxy daunomycin hydrochloride (I-HCl)

To a solution of (X)(0.8 g 1.14 mmol) in dry THF (80 ml) at 0° C. asolution of CH.tbd.CMgBr in THF, (30 ml of 1N solution corresponding toca 30 mol equiv) was added.

After 1 h the mixture was poured into an ice-cold saturated solution ofNH₄ Cl and extracted with methylene chloride. The organic layer waswashed with water, dried over Na₂ SO₄ and evaporated to leave (XI) whichwas dissolved in acetic acid (15 ml) and lead tetraacetate (500 mg 1.12mmol) was added. After 24 h water was added to the mixture to give (XII)as a red precipitate that was filtered and dried.

This precipitate was dissolved in acetone (75 ml) and treated with redHgO (0.6 g, 2.76 mmol) and 7% aqueous H₂ SO₄ (75 ml).

The mixture was heated under reflux for 5' and allowed to cool to roomtemperature. After usual work-up a red solid was obtained (XIII) (0.425g 50% of yield based upon (X).

0.3 g of (XIII) (R₃ ═CF₃ ; R₄ ═--C₆ H₄ NO₂) (0.4 mmol) was dissolved in100 ml of 0.IN NaOH and kept at room temperature for 1 h. The solutionwas brought to pH 8 with 5N HCl and extracted with methylene chloride.

Evaporation of the solvent left a residue that was taken up in 3 ml of amixture of chloroform-methanol 3:2 (v:v). Methanolic 0.IN HCl was addedto adjust the pH to 4.5, after which sufficient diethyl ether was addedto precipitate 0.13 g of the hydrochloride of 4-demethoxy-daunomycin

    ______________________________________                                                (I--HCl)                                                                      mp 183-185° C.                                                         [α].sub.D.sup.20 = +205° (c = 0.1 MeOH)                  ______________________________________                                    

SCHEME ##STR19##

We claim:
 1. A stereospecific process for the preparation of4-demethoxydaunorubicin of formula (I), comprising the stepsof:dissolving the quinone quinizarine epoxide of formula (III):##STR20## in dry toluene, reacting said epoxide at room temperature andfor 24 hours, with a daunosamine of formula II: ##STR21## to give aprotected tetracyclic compound of formula (VIII): ##STR22## having the7(S) configuration, hydrolytically clearing the group in tetrahydrofuranwith 0.1N hydrochloric acid to give a ketone of formula (IX): ##STR23##dissolving said ketone in anhydrous acetonitrile and treating saiddissolved ketone with sodium iodide and trimethylsilyl chloride under anitrogen atmosphere, to obtain the compound of formula (X): ##STR24##treating said compound (X) at a temperature of 0° C., in anhydroustetrahydrofuran with an excess of ethynyl magnesium bromide to give thecompound of formula (XI): ##STR25## treating said compound of formula(XI) with lead tetracetate in acetic acid to obtain the compound offormula (XII): ##STR26## reacting said compound of formula (XII) withred mercuric oxide and aqueous 7% sulphuric acid to give theN,O-protected 4-demethoxydaunorubicin of formula (XIII): ##STR27##hydrolysing said compound of formula (XIII) in 0.1N aqueous sodiumhydroxide, at room temperature, to give the 4-demethoxydaunorubicin offormula (I).
 2. The process of claim 1, further comprising treating the4-demethoxydaunorubicin of formula (I) with anhydrous hydrochloric acidin methanol to form the hydrochloride salt of the4-demethoxydaunorubicin of formula (I).